CN112934282A - Self-feedback high-flux microfluidic system and method - Google Patents
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Abstract
The invention discloses a self-feedback high-throughput microfluidic system and a method, wherein the method comprises the following steps: the system comprises a micro-fluidic chip, a multi-channel micro-fluidic control module, an automatic imaging module and an analysis control system; an analysis control system for: controlling an automatic imaging module to shoot a biological tissue or cell sample cultured in the microfluidic chip to obtain a biological image, analyzing the biological image in real time to obtain an experimental result, and feeding the experimental result back to the multichannel microfluidic control module; and the multi-channel microfluid control module adjusts experimental parameters in the microfluidic chip according to experimental results. The invention combines the advantages of the microfluidic technology, the automatic microscopic imaging technology, the automatic image analysis and the data processing technology, realizes the full-automatic self-feedback high-flux microscopic imaging through integration, can be applied to complex biomedical experiments such as high-flux drug screening based on living cells and biological tissues and the like, and has high accuracy and more convenience.
Description
Technical Field
The invention relates to the technical field of biological sample culture, in particular to a self-feedback high-throughput microfluidic system and a method.
Background
Drug screening is a key step in new drug development, and the discovery of innovative drugs requires screening of a large number of compound samples by adopting appropriate drug action targets, analysis and detection of the safety and effectiveness of drugs are required in preclinical experimental stages, thousands of samples are involved in one process, and condition factors including drug concentration, delivery modes, biological micro-environments, cell types and the like are considered. At present, the most effective combination therapy for clinical anti-tumor needs to screen tens of thousands or hundreds of thousands of drug combination schemes. In the prior art, biological samples under the condition of medicines are detected one by one in a manual mode, the work is boring, the steps are single, manual errors are easily generated in screening results, the time consumption is long, and serious problems that the medicine research and development cost is greatly increased, the treatment is delayed and the like are caused.
The full-automatic operation system replaces manual operation, obviously has a plurality of advantages, the whole experiment process is controlled by a computer through operation software, the operation is simple, the workload of a traditional experiment detection means for months can be finished by a single person in a short time, the screening result is real and reliable, and a plurality of medicines can be screened in a plurality of samples at the same time, so that higher flux is realized. Currently, the technical means for realizing the full-automatic biomedical experiment and having the capability of culturing living cells and biological tissues mainly include a full-automatic robot (e.g., a maga MegaLab biosafety robot) and a valve-controlled microfluidic technology. Compared with the expensive price, huge volume and complex operation program of the biosafety robot, the valve-controlled microfluidic technology is realized based on the microfluidic chip, has the advantages of micro, high efficiency, high flux, automation, low manufacturing cost and the like, and can automatically complete operations such as inoculation of cells, replacement or addition of culture solution, addition and cleaning of medicines, addition of cell staining reagents and the like. In addition, the biological sample and the medicine amount consumed by the micro-fluidic chip are extremely low, the nano-liter level is achieved, and the experiment cost can be greatly saved.
The high-throughput screening based on the micro-fluidic technology at present mainly comprises three main categories: 1. microfluidic screening systems based on droplet patterns. The system utilizes two mutually insoluble liquid drops with cells wrapped by two phases to complete operations such as cell culture, drug stimulation to the cells and the like in the liquid drops. For example: a PDMS channel chip designed by Wong et al for forming cell droplets for drug screening (Wong AH, et al. sci Rep,2017,7, 9109). The microfluidic chip provided by Hefeng screen et al is used for detecting exosome GPC1 protein and application thereof in early diagnosis of pancreatic cancer (CN 111208297A). The system can form a large number of cell drops in a short time, improve the flux of experiments and effectively reduce the consumption of reagents. However, such systems do not allow for the renewal of nutrients in the cultured cells. 2. Microfluidic cell screening systems based on microarray formats. The system generates a cell droplet array on a carrier for cell culture by a certain means, and controls and analyzes the cell droplet array. For example, Lee et al constructed a microarray chip system named hydrogel droplet array for use in high-throughput drug screening experiments (Lee M, et al. A spectral biosensing device (CN 112067585 a) proposed by liu xiao hu et al. Systems of this type are often complex and the implementation of these operations relies on highly accurate and self-contained fluid handling equipment. 3. Microfluidic cell screening systems based on perfusion flow patterns. This type of system inoculates cells in a culture chamber in a microchannel by controlling the continuous flow of fluid through the microchannel and accomplishes drug stimulation of the cells. For example: wang et al designed a microfluidic Chip based on the perfusion flow mode for orthogonalized drug screening (Wang Z, et al lab Chip,2007,7, 740). Yangjinyi et al propose a microfluidic chip immunoassay kit and a detection method thereof (CN 111077319A).
However, at present, most of high-throughput screening systems based on microfluidic technology are difficult to automate the whole process operations of culturing, dosing, detecting, analyzing and the like under ultra-low and high-throughput levels, difficult to automatically realize the addition and stimulation of drugs with different compositions and concentrations on a large scale, and difficult to automatically realize the rapid high-throughput detection of screening results.
Disclosure of Invention
The present invention is directed to solving, at least to some extent, one of the technical problems in the art described above. Therefore, the invention aims to provide a self-feedback high-throughput microfluidic system and a method, and the system combines a microfluidic technology, an automatic microscopic imaging technology, real-time data analysis and a feedback technology to realize the culture of high-throughput cells and biological tissues; controllable medicine proportion (including mixing of multiple medicine components, medicine dilution and the like); the stress response of cells and biological tissues to the medicine is detected and analyzed in real time; and the experimental result is fed back, so that the experimental conditions are adjusted in real time, a dynamic drug environment is constructed, and the efficiency and the accuracy of drug screening are obviously improved.
In order to achieve the above object, the present invention provides a self-feedback high-throughput microfluidic system, comprising: the system comprises a micro-fluidic chip, a multi-channel micro-fluidic control module, an automatic imaging module and an analysis control system; wherein,
the multi-channel micro-fluid control module is connected with the micro-fluid control chip;
the analysis control system is respectively connected with the multi-channel microfluid control module and the automatic imaging module and is used for:
controlling the automatic imaging module to shoot a biological tissue or cell sample cultured in the microfluidic chip to obtain a biological image, analyzing the biological image in real time to obtain an experimental result, and feeding the experimental result back to the multi-channel microfluidic control module;
and the multi-channel microfluidic control module adjusts experimental parameters in the microfluidic chip according to the experimental result.
According to some embodiments of the invention, the multi-channel microfluidic control module comprises: the device comprises an electromagnetic valve, a liquid outlet micro-pipeline, a USB serial port interface, a gas line interface, a power supply interface, a control card, a lead, a gas supply line, a liquid storage pipe, a gas guide rail and a liquid storage pipe frame; wherein,
one end of the USB serial port interface is connected with the analysis control system, and the other end of the USB serial port interface is connected with the control card;
one end of the power interface is connected with the mains supply, and the other end of the power interface is connected with the control card;
one end of the air line interface is connected with the air compressor or the compressed air bottle, and the other end of the air line interface is connected with the control card;
the lead is arranged between the control card and the electromagnetic valve;
the air supply line is used for connecting the air outlet of the electromagnetic valve with the liquid storage pipe;
the liquid storage pipe is arranged on the liquid storage pipe frame; the liquid storage pipe is stored with target liquid;
one end of the liquid outlet micro-pipeline extends into the liquid storage pipe, and the other end of the liquid outlet micro-pipeline is connected with the micro-fluidic chip;
the electromagnetic valve is arranged on the gas path of the gas guide rail;
the control card is respectively connected with the USB serial port interface, the power supply interface and the air line interface; a solenoid valve connection for:
receiving the experimental result obtained by the analysis control system sent by the USB serial port interface;
obtaining a power supply based on the power interface;
acquiring target gas based on the gas line interface, and enabling the target gas to enter the gas guide rail through a gas inlet hole of the gas guide rail;
and controlling the switch of the electromagnetic valve to control the on-off of the gas circuit.
According to some embodiments of the invention, the reservoir comprises: a tube body, an external spiral luer head and a tube cover; wherein,
the liquid outlet micro-pipeline extends into the liquid storage pipe through a round hole arranged on the pipe body and is contacted with the bottom of the liquid storage pipe;
and the outer spiral luer head penetrates through one end of the upper part of the pipe cover and is connected with the air outlet hole of the electromagnetic valve through the air supply wire.
According to some embodiments of the invention, the microfluidic chip comprises: the device comprises a liquid inlet, a micro-flow channel, a first control valve, a second control valve, a third control valve, a culture cavity, a fourth control valve and a liquid outlet; wherein,
the liquid inlet is arranged at the first end part of the micro-flow channel;
the micro-flow channel is sequentially provided with the first control valve, the second control valve, the third control valve and the fourth control valve;
the culture cavity is arranged between the third control valve and the fourth control valve; a biological tissue or cell sample which is marked by fluorescence is cultured in the culture cavity;
the liquid outlet is arranged at the second end part of the micro-flow channel.
According to some embodiments of the invention, the automatic imaging module comprises an excitation light source and an imaging device;
the excitation light source comprises a laser device and an excitation light path, the laser device emits laser through the excitation light path, the wavelength of the laser is matched with the excitation wavelength of the fluorescence-labeled biological tissue or cell sample, and light spots of the light source cover each independent culture cavity;
the imaging device comprises a CCD camera used for collecting biological images and an attached light path used for collecting the biological images, and the biological images are fluorescence images.
According to some embodiments of the invention, the air compressor or the compressed gas cylinder inputs the target gas through the gas line interface at a pressure ranging from 20psi to 50 psi;
the flow speed range of the liquid in the micro-fluidic chip is 0.1mm/s-1 mm/s.
According to some embodiments of the invention, the tube body of the liquid storage tube is made of plastic, and the parameter is 5-15 ml; the diameter of the round hole arranged on the tube body is 1.5 mm; the diameter of the pipe cover is 22mm, a round hole with the diameter of 3.5mm is formed in the center of the pipe cover, the head of the outer spiral luer head with the diameter of 3.2mm extends out from inside to outside and penetrates through the round hole with the diameter of 3.5mm, and the outer spiral luer head is fixed from the inside of the pipe cover by waterproof anti-pressure glue; the length of the air supply line ranges from 20cn to 30 cm.
According to some embodiments of the invention, the microfluidic chip comprises 56 culture chambers, the parameters of the culture chambers are 400 μm long by 400 μm wide by 150 μm high; the parameters of the microfluidic channel are 100 micrometers wide by 25 micrometers high; the parameters of the first control valve, the second control valve, the third control valve and the fourth control valve are 100 micrometers in length and 100 micrometers in width; the microfluidic chip comprises 22 liquid inlets.
According to some embodiments of the invention, the first control valve, the second control valve and the third control valve are opened and closed according to a preset control logic to drive liquid into the culture chamber; when the fourth control valve is opened, the liquid in the culture cavity flows out through the liquid outlet through the fourth control valve.
According to some embodiments of the present invention, a method for self-feedback high-throughput drug screening based on the self-feedback high-throughput microfluidic system described above comprises:
sequentially turning on a power supply of an external computer, turning on a power supply of an automatic imaging module, connecting a power supply interface plug into a power supply interface of a multi-channel microfluid control module, and respectively connecting a control card with the external computer by using a USB serial port interface;
preparing a high-flux drug screening platform, connecting a liquid inlet of a microfluidic chip with a liquid outlet micropipe of a multi-channel microfluidic control module, and opening and closing an electromagnetic valve by connecting a control system in an external computer to pressurize the microfluidic chip so as to realize control on liquid transportation in the microfluidic chip;
initializing an analysis control system, namely starting analysis control software on an external computer provided with the analysis control system, connecting a control card of the automatic imaging module and the multi-channel microfluid control module, and after the connection is successful, automatically initializing the hardware of the automatic imaging module and the hardware of the control card by the analysis control software;
loading a biological tissue or cell sample; introducing a liquid carrying a biological tissue or cell sample into the culture chamber through the microfluidic channel; the loading mode can adopt a mode of opening the culture chambers one by one for introduction, and can open the whole row of culture chambers for one-time introduction;
step five, medicine input control, namely pressurizing and guiding a plurality of medicines to be screened and cleaning solution into the microfluidic chip, controlling peristaltic pumps below the medicine liquid interfaces to circularly switch at different frequencies to mix the input medicines at different proportions or dilute the input medicines at different concentrations, and guiding the input medicines into a specified culture cavity;
step six, arranging an automatic imaging module in analysis control software to perform multi-point tracking shooting, and observing the dynamic change of a test biological sample in the culture cavity;
step seven, setting target state parameters of a test object after drug stimulation, setting sequences and concentrations of added drugs and sequences and concentrations of auxiliary drugs in an analysis control system, starting analysis control software to start an unattended drug screening process, and performing timing comparison and full-automatic drug addition; the target parameters comprise cell number, biological tissue size and fluorescence intensity;
and step eight, automatically selecting the target drugs and the auxiliary drugs in the drug sequences by the analysis control system through real-time comparison, combining various drug concentration stimulation test objects, finishing screening and outputting the drug sequences used when the target is reached, respective concentrations and action time when the set values and the specific time are met, and outputting the final expression results of the biological tissues or cell samples during drug screening.
Has the advantages that:
(1) the self-feedback high-throughput microfluidic system controls the multichannel microfluidic control module to carry out nanoliter precision control on liquid transportation of the microfluidic chip through the analysis control system, simultaneously obtains a biological image of a biological sample through the automatic imaging module, obtains development trends of cells and biological tissues under drug conditions through real-time analysis of the biological image, and feeds an experimental result back to the multichannel microfluidic control module, so that drug conditions in a culture cavity of the microfluidic chip are adjusted in real time, and dynamically adjustable drug screening is realized;
(2) compared with the existing screening system based on a molecular structure, the system can obtain an experimental result which can reflect the effectiveness of drug molecules in a complex life system and can obtain a drug screening result meeting the clinical application requirement more quickly and accurately;
(3) the self-feedback high-throughput microfluidic system integrates an analysis control system, a multi-channel microfluidic control module, a microfluidic chip and an automatic imaging module into an automatic integrated system, and overcomes the defects of independence, high manual participation degree, long time consumption, easy larger system error and the like of the conventional drug screening technology.
Additional features and advantages of the invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objectives and other advantages of the invention will be realized and attained by the structure particularly pointed out in the written description and drawings.
The technical solution of the present invention is further described in detail by the accompanying drawings and embodiments.
Drawings
The accompanying drawings, which are included to provide a further understanding of the invention and are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the principles of the invention and not to limit the invention. In the drawings:
FIG. 1 is a schematic diagram of a self-feedback, high-throughput microfluidic system according to one embodiment of the present invention;
FIG. 2 is a schematic connection diagram of a microfluidic chip according to an embodiment of the present invention;
fig. 3 is a schematic diagram of different colors of food colors in a microfluidic chip demonstrating different drug environmental conditions, according to one embodiment of the present invention.
Reference numerals:
1-an analytical control system; 2-a multi-channel microfluidic control module; 201-electromagnetic valve; 202-liquid outlet micro-pipeline; 203-USB serial port interface; 204-gas line interface; 205-power interface; 206-a control card; 207-a wire; 208-air outlet holes; 209-air inlet; 210-an air supply line; 211-a liquid storage tube; 212-a tube body; 213-external spiral luer head; 214-a tube cover; 215-air guide rail, 216-liquid storage tube rack; 3-a microfluidic chip; 4-an automatic imaging module; 301-liquid inlet; 302-a microfluidic channel; 303-a first control valve; 304-a second control valve; 305-a third control valve; 306-a culture chamber; 307-a fourth control valve; 308-liquid outlet.
Detailed Description
The preferred embodiments of the present invention will be described in conjunction with the accompanying drawings, and it will be understood that they are described herein for the purpose of illustration and explanation and not limitation.
As shown in fig. 1, an embodiment of the invention provides a self-feedback high-throughput microfluidic system, including: the system comprises a micro-fluidic chip 3, a multi-channel micro-fluidic control module 2, an automatic imaging module 4 and an analysis control system 1; wherein,
the multi-channel micro-fluid control module 2 is connected with the micro-fluid control chip 3;
the analysis control system 1 is respectively connected with the multi-channel microfluid control module 2 and the automatic imaging module 4, and is configured to:
controlling the automatic imaging module 4 to shoot a biological tissue or cell sample cultured in the microfluidic chip 3 to obtain a biological image, analyzing the biological image in real time to obtain an experimental result, and feeding the experimental result back to the multi-channel microfluidic control module 2;
and the multi-channel micro-fluid control module 2 adjusts the experimental parameters in the micro-fluid control chip 3 according to the experimental result.
The working principle of the technical scheme is as follows: the analysis control system 1 is an external computer (PC) installed with relevant software, and includes relevant imaging software, relevant control software, and relevant analysis software. The analysis control system 1 is respectively connected with the multi-channel microfluid control module 2 and the automatic imaging module 4, the control of liquid transportation in the microfluidic chip 3 is realized by controlling the multi-channel microfluid control module 2, the automatic imaging module 4 is controlled to acquire image signals, namely, the automatic imaging module 4 is controlled to shoot biological tissues or cell samples cultured in the microfluidic chip 3, a biological image is acquired, the biological image is analyzed in real time to acquire an experimental result, the experimental result is fed back to the multi-channel microfluid control module 2, and then the experimental conditions in the microfluidic chip 3 are adjusted. The micro-fluidic chip 3 is connected with the multi-channel micro-fluidic control module 2, so that high-flux cell and tissue culture can be realized, and liquid can be changed under the control of the multi-channel micro-fluidic. The automatic imaging module 4 is connected to the analysis control system 1, is controlled by the analysis control system 1, performs timing and fixed-point imaging at a designated position (positioning culture chamber 306 and liquid input port), and transmits the acquired biological image to the analysis control system 1.
The beneficial effects of the above technical scheme are that: the microfluidic technology, the automatic microscopic imaging technology, the real-time data analysis and the feedback technology are combined to realize the culture of high-throughput cells and biological tissues; controllable medicine proportion (including mixing of multiple medicine components, medicine dilution and the like); the stress response of cells and biological tissues to the medicine is detected and analyzed in real time; and the experimental result is fed back, so that the experimental conditions are adjusted in real time, a dynamic drug environment is constructed, and the efficiency and the accuracy of drug screening are obviously improved.
According to some embodiments of the invention, the multi-channel microfluidic control module 2 comprises: the device comprises an electromagnetic valve 201, a liquid outlet micro-pipeline 202, a USB serial port interface 203, a gas line interface 204, a power supply interface 205, a control card 206, a conducting wire 207, a gas supply line 210, a liquid storage pipe 211, a gas conducting rail 215 and a liquid storage pipe frame 216; wherein,
one end of the USB serial port interface 203 is connected with the analysis control system 1, and the other end is connected with the control card 206;
one end of the power interface 205 is connected to the mains supply, and the other end is connected to the control card 206;
one end of the air line interface 204 is connected with the air compressor or the compressed air bottle, and the other end is connected with the control card 206;
the lead 207 is arranged between the control card 206 and the electromagnetic valve 201;
the air supply line 210 is used for connecting the air outlet hole 208 of the electromagnetic valve 201 with the liquid storage pipe 211;
the liquid storage pipe 211 is arranged on the liquid storage pipe frame 216; the liquid storage pipe 211 stores target liquid;
one end of the liquid outlet micro-pipeline 202 extends into the liquid storage tube 211, and the other end is connected with the micro-fluidic chip 3;
the electromagnetic valve 201 is arranged on the gas path of the gas guide rail 215;
the control card 206 is respectively connected with the USB serial port interface 203, the power supply interface 205 and the air line interface 204; the solenoid valve 201 is connected to:
receiving the experimental result obtained by the analysis control system 1 sent by the USB serial port interface 203;
obtaining power based on the power interface 205;
acquiring target gas based on the gas line interface 204, and enabling the target gas to enter the air guide rail 215 through the air inlet hole 209 of the air guide rail 215;
and controlling the switch of the electromagnetic valve 201 to control the on-off of the air path.
The working principle of the technical scheme is as follows: the USB serial port interface 203, the air line interface 204, the power supply interface 205 and the electromagnetic valve 201 are all connected with a control card 206. The control card 206 is connected to the analysis control system 1, receives signals of the analysis control system 1, includes experimental results, and is controlled by the analysis control system 1; the control card 206 is connected with the analysis control system 1 through the USB serial port interface 203, is connected with the commercial power through the power interface 205, and is connected with the air compressor or the compressed air bottle through the air line interface 204. The control card 206 is connected with the electromagnetic valve 201 through a wire 207; the target gas is connected into the air guide rail 215 through the air inlet 209 of the air guide rail 215, as shown in fig. 1, the inlet air can be divided into 8 paths to be discharged, and each path is connected with one electromagnetic valve 201 for control; the solenoid valve 201 outputs the target gas through a gas outlet 208 of the solenoid valve 201, and is connected to a reservoir 211 by a gas supply line 210. The control card 206 controls the electromagnetic valve 201 to open and close the air passage, and the opening and closing signal of the electromagnetic valve 201 is provided by the control card 206, namely, the power supply is conducted and the power is cut off. The air inlet and outlet hole 208 of the electromagnetic valve 201 is matched with the air outlet hole 208 on the air guide rail 215. The control card 206 receives signals of the analysis control system 1 through communication of the USB serial port interface 203. The on/off of the electromagnetic valve 201 is controlled by the communication of the USB serial port interface 203. The opening and closing of each solenoid valve 201 can be controlled by software of the analysis control system 1. The solenoid valve 201 can control the output of the liquid in the liquid storage pipe 211 through a gas switch. The tube body 212 of the liquid storage tube 211 is a plastic tube, and the tube body 212 stores liquid to be injected into the microfluidic chip 3. The tube body 212 of the liquid storage tube 211 is provided with a round hole, and the liquid micro-pipeline penetrates through the round hole and contacts with the tube bottom of the liquid storage tube 211. When the pressure in the closed pipe body 212 increases, the liquid flows out along the liquid outlet micro-pipeline 202. The cap 214 of the liquid storage tube 211 is matched with the tube body 212 to form an airtight environment of the liquid storage tube 211. The pipe cover 214 is provided with an external spiral luer head 213, the head part of the external spiral luer head 213 extending out of the pipe cover 214 is connected with the air outlet hole 208 corresponding to the electromagnetic valve 201 through the air supply line 210, the electromagnetic valve 201 can control the pressure of the liquid storage pipe 211 through controlling the air circuit switch, and further control the liquid in the liquid storage pipe 211 to flow out through the liquid outlet micro-pipeline 202. The liquid storage pipe 211 is arranged on the liquid storage pipe frame 216 after being combined, and is stable and portable. The control process of the multi-channel microfluid control module 2 is as follows: the control card 206 receives the control signal of the analysis control system 1 through the USB serial port interface 203; the control card 206 controls the on-off of the electromagnetic valve 201 through a lead 207; the electromagnetic valve 201 controls the pressure of the liquid storage pipe 211 by controlling the air circuit switch, and further controls the outflow of the liquid in the liquid storage pipe 211 through the liquid outlet micro-pipeline 202. The liquid outlet micro-pipeline 202 is connected with the micro-fluidic chip 3, and liquid output by the liquid outlet micro-pipeline 202 enters the micro-fluidic chip 3 through the liquid inlet 301. The power interface 205 is a DC power plug. Due to the control of the control card 206, the gas pressure drives the liquid to make the control valve channel in the micro-fluidic chip 3 generate spatial deformation, thereby realizing the control of the fluid layer micro-channel switch in the micro-fluidic chip 3.
The beneficial effects of the above technical scheme are that: the liquid stored in the liquid storage tube 211 is intelligently input into the microfluidic chip 3, the air pressure entering the liquid storage tube 211 is intelligently controlled, and the flow speed of the liquid is further controlled.
According to some embodiments of the invention, the reservoir 211 comprises: a tube body 212, an external spiral luer head 213 and a tube cover 214; wherein,
the liquid outlet micro-pipeline 202 extends into the liquid storage pipe 211 through a round hole arranged on the pipe body 212 and is contacted with the bottom of the liquid storage pipe 211;
the outer spiral luer 213 passes through one end of the upper part of the pipe cover 214 and is connected with the air outlet hole 208 of the solenoid valve 201 through the air supply wire 210.
According to some embodiments of the invention, the microfluidic chip 3 comprises: a liquid inlet 301, a micro-flow channel 302, a first control valve 303, a second control valve 304, a third control valve 305, a culture cavity 306, a fourth control valve 307 and a liquid outlet 308; wherein,
the liquid inlet 301 is arranged at a first end part of the microfluidic channel 302;
the microfluidic channel 302 is sequentially provided with the first control valve 303, the second control valve 304, the third control valve 305 and the fourth control valve 307;
the culture chamber 306 is arranged between the third control valve 305 and the fourth control valve 307; a biological tissue or cell sample which is marked by fluorescence is cultured in the culture cavity 306;
the liquid outlet 308 is disposed at a second end of the micro-flow channel 302.
Liquid output by the multi-channel micro-fluid control module 2 through the liquid outlet micro-pipeline 202 enters the micro-fluid control chip 3 through the liquid inlet 301; the liquid is a medicine containing different concentrations and different components; when the first control valve 303, the second control valve 304, the third control valve 305 and the fourth control valve 307 are all opened, the injected liquid flows into the culture chamber 306 through the microfluidic channel 302 to act on the cultured biological sample. The liquid inlet end of the micro-fluidic chip 3 is controlled by a liquid driving pump (peristaltic pump) consisting of a first control valve 303, a second control valve 304 and a third control valve 305, the first control valve 303, the second control valve 304 and the third control valve 305 sequentially control the micro-valve switches through the control logic of (100, 110, 010, 011, 001) to drive the liquid to flow, wherein 1 represents that the control valve is opened, the liquid can flow in, 0 represents that the control valve is closed, and the liquid cannot flow in. The liquid outlet end of the microfluidic chip 3 is controlled by the fourth control valve 307, and when the fourth control valve 307 is opened, the liquid can flow out through the liquid outlet 308 via the fourth control valve 307.
According to some embodiments of the present invention, the automatic imaging module 4 comprises an excitation light source and an imaging device;
the excitation light source comprises a laser device and an excitation light path, the laser device emits laser through the excitation light path, the wavelength of the laser is matched with the excitation wavelength of the fluorescence-labeled biological tissue or cell sample, and light spots of the light source cover each independent culture cavity 306;
the imaging device comprises a CCD (charge coupled device) camera for collecting biological images and an attached light path for collecting the biological images, and the biological images are fluorescence images.
The beneficial effects of the above technical scheme are that: the accuracy of the acquired biological image is improved, the accuracy of biological image analysis is improved conveniently, and further more suitable experimental conditions are acquired and adjusted.
According to some embodiments of the present invention, the air compressor or compressed gas cylinder inputs the target gas through the gas line interface 204 at a pressure ranging from 20psi to 50 psi;
the flow speed range of the liquid in the micro-fluidic chip 3 is 0.1mm/s-1 mm/s.
The working principle and the beneficial effects of the technical scheme are as follows: the target gas may be air or a safe gas such as nitrogen, and is used to adjust the air pressure in the liquid storage tube 211, thereby realizing the liquid transportation with controllable flow rate and flow rate in the microfluidic chip 3. The accuracy of liquid flow velocity and flow control is improved, effective absorption of liquid in the culture cavity 306 is guaranteed, and the culture efficiency of biological samples is improved.
According to some embodiments of the present invention, the tube body 212 of the liquid storage tube 211 is made of plastic, and the parameters are 5-15 ml; the diameter of the round hole arranged on the tube body 212 is 1.5 mm; the diameter of the pipe cover 214 is 22mm, a round hole of 3.5mm is arranged in the center of the pipe cover 214, the head of the outer spiral luer head 213 with the diameter of 3.2mm extends from inside to outside, penetrates through the round hole of 3.5mm, and the outer spiral luer head 213 is fixed from the inside of the pipe cover 214 by waterproof and pressure-resistant glue; the length of the gas supply line 210 ranges from 20cn to 30 cm.
According to some embodiments of the present invention, the microfluidic chip 3 comprises 56 culture chambers 306, and the parameters of the culture chambers 306 are 400 μm long × 400 μm wide × 150 μm high; the microfluidic channel 302 has a width of 100 μm × a height of 25 μm; the parameters of the first control valve 303, the second control valve 304, the third control valve 305 and the fourth control valve 307 are 100 μm in length × 100 μm in width; the microfluidic chip 3 comprises 22 liquid inlets 301.
The microfluidic chip 3 comprises 56 independent cell and biological tissue culture chambers 306, each culture chamber 306 is controlled by four control valves, and can culture tissues and cell samples and maintain independently controllable environments with different drug concentrations and combined drugs. 30656 culture cavities, which can be independently controlled or serially controlled, are provided, experimental samples are cultured in the culture cavities, and culture media and liquid containing related medicines injected into the culture cavities 306 are controlled by a multi-channel microfluidic control device.
The culture chamber 306 contains cell lines, primary cells, organoids, and biological tissue micro-blocks. During the living cell experiment, the morphological behavior of cells and cell nucleus can be observed through bright field microscopic imaging, and the sample can be stained and tracked through fluorescent imaging through a living cell stain (such as DAPI, 4', 6-diamidino-2-phenylindole which is a DNA fluorescent marker). Morphological features of organoids and biological tissues are observed primarily by bright field imaging. For the intracellular biomolecule interaction, living cell immunofluorescence staining or transient transfer is adopted for labeling staining, and then fluorescence imaging is used for observation.
The liquid in reservoir 211 is a liquid containing the relevant drug, and refers to different concentrations of a single drug, as well as different drug combinations. The medicine mixing and diluting are completed by controlling the liquid flow in the microfluidic chip 3 by a control valve.
The liquid inlet 301 is connected to the multi-channel microfluidic control module 2, and when the first control valve 303, the second control valve 304, the third control valve 305 and the fourth control valve 307 are all opened, the injected liquid is controlled by the multi-channel microfluidic control module 2. A microfluidic channel 302 (width 100 μm × height 25 μm) is correspondingly connected below the liquid inlet 301, the first control valve 303, the second control valve 304, the third control valve 305, and the fourth control valve 307 are all opened, and the liquid injected into the liquid inlet 301 flows through the culture chamber 306 along the microfluidic channel 302.
According to some embodiments of the present invention, the first control valve 303, the second control valve 304, and the third control valve 305 open and close the valves according to a preset control logic to drive the liquid into the culture chamber 306; when the fourth control valve 307 is opened, the liquid in the culture chamber 306 flows out through the fourth control valve 307 via the liquid outlet 308.
According to some embodiments of the present invention, a method for self-feedback high-throughput drug screening based on the self-feedback high-throughput microfluidic system described above comprises:
sequentially turning on a power supply of an external computer, turning on a power supply of the automatic imaging module 4, inserting a power supply interface 205 plug into a power supply interface 205 of the multi-channel microfluid control module 2, and respectively connecting a control card 206 with the external computer by using a USB serial port interface 203;
preparing a high-flux drug screening platform, connecting a liquid inlet 301 of the microfluidic chip 3 with a liquid outlet micro-pipeline 202 of the multi-channel microfluidic control module 2, and opening and closing an electromagnetic valve 201 by connecting a control system in an external computer to pressurize the microfluidic chip 3 so as to realize control on liquid transportation in the microfluidic chip 3;
step three, initializing the analysis control system 1, namely starting analysis control software on an external computer provided with the analysis control system 1, connecting the automatic imaging module 4 with the control card 206 of the multi-channel microfluid control module 2, and after the connection is successful, automatically initializing the hardware of the automatic imaging module 4 and the hardware of the control card 206 by the analysis control software;
loading a biological tissue or cell sample; introducing a fluid carrying a biological tissue or cell sample into a culture chamber 306 through a microfluidic channel 302; the loading mode can adopt a mode of opening the culture cavities 306 one by one for introduction, and can open the whole row of culture cavities 306 for introduction at one time;
fifthly, medicine input control, namely pressurizing and guiding a plurality of medicines to be screened and cleaning solution into the microfluidic chip 3, controlling peristaltic pumps below the medicine liquid interfaces to circularly switch at different frequencies to mix the input medicines at different proportions or dilute the input medicines at different concentrations, and guiding the input medicines into a specified culture cavity 306;
step six, in the analysis control software, the automatic imaging module 4 is arranged to perform multi-point tracking shooting, and the dynamic change of the test biological sample in the culture cavity 306 is observed;
step seven, setting target state parameters of a test object after drug stimulation, setting sequences and concentrations of added drugs and sequences and concentrations of auxiliary drugs in the analysis control system 1, starting analysis control software to start an unattended drug screening process, and performing timing comparison and full-automatic drug addition; the target parameters comprise cell number, biological tissue size and fluorescence intensity;
and step eight, through real-time comparison, the analysis control system 1 automatically selects a target drug and an auxiliary drug in the drug sequence, combines various drug concentration stimulation test objects, finishes screening and outputs the drug sequence, respective concentration and action time used when the target is reached and outputs the final expression result of the biological tissue or cell sample during drug screening when the set value and specific time are met.
In one embodiment, the number of the electromagnetic valves 201 is 1-24, and each electromagnetic valve 201 is connected with the control card 206 through a wire 207; the air inlet line is connected to the air inlet 209 of each air guide rail 215, and each air guide rail 215 comprises 8 air outlets; each solenoid valve 201 corresponds to an air outlet 208 on the air guide rail 215 and is connected with a corresponding liquid storage pipe 211 through an air line.
In a specific embodiment, S1, the system is connected as shown in fig. 1, and the external computer, the automatic imaging module 4 and the multi-channel microfluidic control module 2 are powered on; s2, starting a software system, manually connecting the automatic imaging module 4 and the control card 206 of the multi-channel microfluid control module 2 respectively, and automatically initializing the system after successful connection; s3, connecting the microfluidic chip 3 shown in the figure 3 into a system, wherein the connection completion effect is as shown in figure 2, and the liquid outlet micro-pipeline 202 of the multi-channel microfluidic control module 2 is connected with a control valve of the microfluidic chip 3 to control the delivery of liquid (different medicines); a liquid inlet 301 of the microfluidic chip 3 is connected with a drug inlet and is driven by gas pressure, and then the connected chip is placed on a microscope stage; culturing living cells, organoids or biological tissues to be detected in the culture cavity 306; to achieve fluorescence microscopy, the biological sample is stained with live cells prior to introduction into the chip; in order to observe the dynamic response process of intracellular signal pathways and protein molecule interaction, a specific intracellular protein molecule is labeled by using a fluorescent small molecule or transient transfection; s4, setting the driving air pressure for conveying liquid carrying different medicines to 1.5psi through a digital pressure reducing valve, connecting the driving air pressure to a liquid inlet 301 of a microfluidic chip 3 through a liquid outlet micro-pipeline 202, further controlling the input of the medicines by opening and closing a micro-flow valve controlled by a multi-channel microfluidic control module 2, and enabling the input medicines to be mixed in different proportions or diluted in concentration by controlling peristaltic pumps below a medicine liquid interface to circulate on and off at different frequencies and leading the input medicines into a specified culture cavity 306; s5, switching the objective lens of the automatic imaging module 4 to 20X, then moving the objective table to the culture cavity 306 of the microfluidic chip 3, adjusting the focal length of the objective lens, recording the positions of the upper left culture cavity 306 and the lower right culture cavity 306, automatically calculating the positions of other culture cavities 306 in the matrix by a program, and generating a position array; s6, setting the expected value of the effect of the screened drug on the biological sample, including the influence on the movement capacity, proliferation rate and death rate of the tumor cells, which are expressed in percentage.
Clicking a start key on an operation interface to start a drug screening process taking the specific cell number as a target, wherein the specific process comprises the following steps:
a) the software controls the control card 206 of the multi-channel microfluid control module 2 through a USB serial port line to cut off the power supply of the electromagnetic valve 201 of the liquid inlet 301 control valve, so that the target medicine combination (subjected to automatic dilution and medicine mixing) enters the culture cavity 306 filled with living cells;
b) the software-controlled automatic imaging module 4 acquires bright field and fluorescence images in the culture cavity 306, then transmits the bright field and fluorescence images to the analysis control system 1, the analysis control system 1 identifies single cells in the images, performs single cell tracking between frames, and further collects the morphology and behavior of the single cells under the drug environment condition and intracellular biochemical reaction information;
c) the experimental result is fed back to the multi-channel microfluidic control module 2 by real-time calculation of the analysis control system 1, so that the experimental conditions in the microfluidic chip 3 are controlled. For example, when the number of tumor cell deaths is less than the set target under single drug conditions, the system selects to increase the amount of perfusion drug in a specific culture chamber 306, and simultaneously add an auxiliary drug causing tumor cell death in another culture chamber 306, and continuously observe the stress response of cells and biological tissues under dynamic drug conditions. Stopping perfusion of the target drug into the culture chamber 306 when the set target is reached, or replacing another drug according to the setting; the software sends out an instruction to supply power to the target electromagnetic valve 201 to control the opening and closing of different valves around the culture cavity 306 and the liquid inlet 301;
d) the operation process of constructing different drug environments (single drug concentration gradient and combined drug) in different culture cavities 306 and detecting the effectiveness of the drugs based on living cells is similar to step c;
e) when the effect of multiple drugs is researched, the software system can automatically take the effect of one drug on cells as an initial input parameter, then adjust the drug concentration and add auxiliary drugs according to the experimental result, and finally realize the screening of the effect of the combined drug of multiple drug components and the dynamic drug concentration on the cells. Or, combining ten thousands of high-throughput chips, directly constructing drug combinations with different concentrations in the independent culture cavity 306, and screening out the optimal drug combination by observing morphological behaviors of cells;
f) and evaluating the synergy and inhibition effect among the medicine components by adopting a decision tree machine learning method.
Based on the beneficial effect of self-feedback high-throughput microfluidic system: (1) the self-feedback high-flux microfluidic system controls the multi-channel microfluidic control module 2 to carry out nano-liter precision control on liquid transportation of the microfluidic chip 3 through the analysis control system 1, simultaneously obtains a biological image of a biological sample through the automatic imaging module 4, obtains development trends of cells and biological tissues under a drug condition through real-time analysis of the biological image, and feeds an experimental result back to the multi-channel microfluidic control module 2, so that the drug condition in the culture cavity 306 of the microfluidic chip 3 is adjusted in real time, and dynamic adjustable drug screening is realized; (2) compared with the existing screening system based on a molecular structure, the system can obtain an experimental result which can reflect the effectiveness of drug molecules in a complex life system and can obtain a drug screening result meeting the clinical application requirement more quickly and accurately; (3) the self-feedback high-throughput microfluidic system integrates the analysis control system 1, the multi-channel microfluidic control module 2, the microfluidic chip 3 and the automatic imaging module 4 into an automatic integrated system, and overcomes the defects of independence, high manual participation degree, long time consumption, easy larger system error and the like of the conventional drug screening technology.
As will be appreciated by one skilled in the art, embodiments of the present invention may be provided as a method, system, or computer program product. Accordingly, the present invention may take the form of an entirely hardware embodiment, an entirely software embodiment or an embodiment combining software and hardware aspects. Furthermore, the present invention may take the form of a computer program product embodied on one or more computer-usable storage media (including, but not limited to, disk storage, optical storage, and the like) having computer-usable program code embodied therein.
The present invention is described with reference to flowchart illustrations and/or block diagrams of methods, apparatus (systems), and computer program products according to embodiments of the invention. It will be understood that each flow and/or block of the flow diagrams and/or block diagrams, and combinations of flows and/or blocks in the flow diagrams and/or block diagrams, can be implemented by computer program instructions. These computer program instructions may be provided to a processor of a general purpose computer, special purpose computer, embedded processor, or other programmable data processing apparatus to produce a machine, such that the instructions, which execute via the processor of the computer or other programmable data processing apparatus, create means for implementing the functions specified in the flowchart flow or flows and/or block diagram block or blocks.
These computer program instructions may also be stored in a computer-readable memory that can direct a computer or other programmable data processing apparatus to function in a particular manner, such that the instructions stored in the computer-readable memory produce an article of manufacture including instruction means which implement the function specified in the flowchart flow or flows and/or block diagram block or blocks.
These computer program instructions may also be loaded onto a computer or other programmable data processing apparatus to cause a series of operational steps to be performed on the computer or other programmable apparatus to produce a computer implemented process such that the instructions which execute on the computer or other programmable apparatus provide steps for implementing the functions specified in the flowchart flow or flows and/or block diagram block or blocks.
It will be apparent to those skilled in the art that various changes and modifications may be made in the present invention without departing from the spirit and scope of the invention. Thus, if such modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include such modifications and variations.
Claims (10)
1. A self-feedback, high-throughput microfluidic system, comprising: the system comprises a micro-fluidic chip, a multi-channel micro-fluidic control module, an automatic imaging module and an analysis control system; wherein,
the multi-channel micro-fluid control module is connected with the micro-fluid control chip;
the analysis control system is respectively connected with the multi-channel microfluid control module and the automatic imaging module and is used for:
controlling the automatic imaging module to shoot a biological tissue or cell sample cultured in the microfluidic chip to obtain a biological image, analyzing the biological image in real time to obtain an experimental result, and feeding the experimental result back to the multi-channel microfluidic control module;
and the multi-channel microfluidic control module adjusts experimental parameters in the microfluidic chip according to the experimental result.
2. The self-feedback, high-throughput microfluidic system of claim 1, wherein said multi-channel microfluidic control module comprises: the device comprises an electromagnetic valve, a liquid outlet micro-pipeline, a USB serial port interface, a gas line interface, a power supply interface, a control card, a lead, a gas supply line, a liquid storage pipe, a gas guide rail and a liquid storage pipe frame; wherein,
one end of the USB serial port interface is connected with the analysis control system, and the other end of the USB serial port interface is connected with the control card;
one end of the power interface is connected with the mains supply, and the other end of the power interface is connected with the control card;
one end of the air line interface is connected with the air compressor or the compressed air bottle, and the other end of the air line interface is connected with the control card;
the lead is arranged between the control card and the electromagnetic valve;
the air supply line is used for connecting the air outlet of the electromagnetic valve with the liquid storage pipe;
the liquid storage pipe is arranged on the liquid storage pipe frame; the liquid storage pipe is stored with target liquid;
one end of the liquid outlet micro-pipeline extends into the liquid storage pipe, and the other end of the liquid outlet micro-pipeline is connected with the micro-fluidic chip;
the electromagnetic valve is arranged on the gas path of the gas guide rail;
the control card is respectively connected with the USB serial port interface, the power supply interface and the air line interface; a solenoid valve connection for:
receiving the experimental result obtained by the analysis control system sent by the USB serial port interface;
obtaining a power supply based on the power interface;
acquiring target gas based on the gas line interface, and enabling the target gas to enter the gas guide rail through a gas inlet hole of the gas guide rail;
and controlling the switch of the electromagnetic valve to control the on-off of the gas circuit.
3. The self-feedback, high-throughput microfluidic system of claim 2, wherein said reservoir comprises: a tube body, an external spiral luer head and a tube cover; wherein,
the liquid outlet micro-pipeline extends into the liquid storage pipe through a round hole arranged on the pipe body and is contacted with the bottom of the liquid storage pipe;
and the outer spiral luer head penetrates through one end of the upper part of the pipe cover and is connected with the air outlet hole of the electromagnetic valve through the air supply wire.
4. The self-feedback, high-throughput microfluidic system of claim 3, wherein said microfluidic chip comprises: the device comprises a liquid inlet, a micro-flow channel, a first control valve, a second control valve, a third control valve, a culture cavity, a fourth control valve and a liquid outlet; wherein,
the liquid inlet is arranged at the first end part of the micro-flow channel;
the micro-flow channel is sequentially provided with the first control valve, the second control valve, the third control valve and the fourth control valve;
the culture cavity is arranged between the third control valve and the fourth control valve; a biological tissue or cell sample which is marked by fluorescence is cultured in the culture cavity;
the liquid outlet is arranged at the second end part of the micro-flow channel.
5. The self-feedback high-throughput microfluidic system of claim 1, wherein said automatic imaging module comprises an excitation light source and an imaging device;
the excitation light source comprises a laser device and an excitation light path, the laser device emits laser through the excitation light path, the wavelength of the laser is matched with the excitation wavelength of the fluorescence-labeled biological tissue or cell sample, and light spots of the light source cover each independent culture cavity;
the imaging device comprises a CCD camera used for collecting biological images and an attached light path used for collecting the biological images, and the biological images are fluorescence images.
6. The self-feedback high-throughput microfluidic system according to claim 2, wherein the air pressure range of the target gas input by the air compressor or the compressed gas cylinder through the air line interface is 20psi-50 psi;
the flow speed range of the liquid in the micro-fluidic chip is 0.1mm/s-1 mm/s.
7. The self-feedback high-throughput microfluidic system according to claim 3, wherein the tube body of said reservoir tube is made of plastic with parameters of 5-15 ml; the diameter of the round hole arranged on the tube body is 1.5 mm; the diameter of the pipe cover is 22mm, a round hole with the diameter of 3.5mm is formed in the center of the pipe cover, the head of the outer spiral luer head with the diameter of 3.2mm extends out from inside to outside and penetrates through the round hole with the diameter of 3.5mm, and the outer spiral luer head is fixed from the inside of the pipe cover by waterproof anti-pressure glue; the length of the air supply line ranges from 20cn to 30 cm.
8. The self-feedback high-throughput microfluidic system according to claim 4, wherein said microfluidic chip comprises 56 culture chambers, and the parameters of said culture chambers are 400 μm in length, 400 μm in width, and 150 μm in height; the parameters of the microfluidic channel are 100 micrometers wide by 25 micrometers high; the parameters of the first control valve, the second control valve, the third control valve and the fourth control valve are 100 micrometers in length and 100 micrometers in width; the microfluidic chip comprises 22 liquid inlets.
9. The self-feedback high-throughput microfluidic system of claim 4, wherein the first control valve, the second control valve and the third control valve are opened and closed according to a preset control logic to drive liquid into the culture chamber; when the fourth control valve is opened, the liquid in the culture cavity flows out through the liquid outlet through the fourth control valve.
10. The method for self-feedback high-throughput drug screening according to any one of claims 1-9, comprising:
sequentially turning on a power supply of an external computer, turning on a power supply of an automatic imaging module, connecting a power supply interface plug into a power supply interface of a multi-channel microfluid control module, and respectively connecting a control card with the external computer by using a USB serial port interface;
preparing a high-flux drug screening platform, connecting a liquid inlet of a microfluidic chip with a liquid outlet micropipe of a multi-channel microfluidic control module, and opening and closing an electromagnetic valve by connecting a control system in an external computer to pressurize the microfluidic chip so as to realize control on liquid transportation in the microfluidic chip;
initializing an analysis control system, namely starting analysis control software on an external computer provided with the analysis control system, connecting a control card of the automatic imaging module and the multi-channel microfluid control module, and after the connection is successful, automatically initializing the hardware of the automatic imaging module and the hardware of the control card by the analysis control software;
loading a biological tissue or cell sample; introducing a liquid carrying a biological tissue or cell sample into the culture chamber through the microfluidic channel; the loading mode can adopt a mode of opening the culture chambers one by one for introduction, and can open the whole row of culture chambers for one-time introduction;
step five, medicine input control, namely pressurizing and guiding a plurality of medicines to be screened and cleaning solution into the microfluidic chip, controlling peristaltic pumps below the medicine liquid interfaces to circularly switch at different frequencies to mix the input medicines at different proportions or dilute the input medicines at different concentrations, and guiding the input medicines into a specified culture cavity;
step six, arranging an automatic imaging module in analysis control software to perform multi-point tracking shooting, and observing the dynamic change of a test biological sample in the culture cavity;
step seven, setting target state parameters of a test object after drug stimulation, setting sequences and concentrations of added drugs and sequences and concentrations of auxiliary drugs in an analysis control system, starting analysis control software to start an unattended drug screening process, and performing timing comparison and full-automatic drug addition; the target parameters comprise cell number, biological tissue size and fluorescence intensity;
and step eight, automatically selecting the target drugs and the auxiliary drugs in the drug sequences by the analysis control system through real-time comparison, combining various drug concentration stimulation test objects, finishing screening and outputting the drug sequences used when the target is reached, respective concentrations and action time when the set values and the specific time are met, and outputting the final expression results of the biological tissues or cell samples during drug screening.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114471762A (en) * | 2022-02-22 | 2022-05-13 | 西北大学 | Design and implementation method of micro-fluidic chip capable of controlling ultralow flow velocity |
| CN114512179A (en) * | 2022-02-02 | 2022-05-17 | 上海图灵智算量子科技有限公司 | Quantum mutual learning mechanism-based binding energy prediction method |
| AU2022279500B1 (en) * | 2022-12-01 | 2023-11-23 | Provectus IP Pty Ltd | Automated cell analyser and method |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103257213A (en) * | 2012-02-20 | 2013-08-21 | 中国科学院大连化学物理研究所 | Totally-integrated high-flux cell level micro-fluidic chip medicine evaluating system |
| US20150017092A1 (en) * | 2012-03-15 | 2015-01-15 | The Regents Of The University Of California | Devices and methods for determining sensitivity to radiation |
| CN105032512A (en) * | 2015-08-25 | 2015-11-11 | 辽宁中医药大学 | Integrated micro-fluidic chip for screening drug compatibility, as well as preparation method and application of integrated micro-fluidic chip |
| CN105084306A (en) * | 2015-07-30 | 2015-11-25 | 西北大学 | Controllable preparation method of large-area laminated micro-nano composite structure |
| CN106471508A (en) * | 2014-06-20 | 2017-03-01 | 康涅狄格儿童医疗中心 | Automated cell culture system and corresponding method |
| CN108679448A (en) * | 2018-06-02 | 2018-10-19 | 哈尔滨工业大学 | Microfluid flow on-line control device and detection method |
-
2021
- 2021-03-26 CN CN202110326904.3A patent/CN112934282A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103257213A (en) * | 2012-02-20 | 2013-08-21 | 中国科学院大连化学物理研究所 | Totally-integrated high-flux cell level micro-fluidic chip medicine evaluating system |
| US20150017092A1 (en) * | 2012-03-15 | 2015-01-15 | The Regents Of The University Of California | Devices and methods for determining sensitivity to radiation |
| CN106471508A (en) * | 2014-06-20 | 2017-03-01 | 康涅狄格儿童医疗中心 | Automated cell culture system and corresponding method |
| CN105084306A (en) * | 2015-07-30 | 2015-11-25 | 西北大学 | Controllable preparation method of large-area laminated micro-nano composite structure |
| CN105032512A (en) * | 2015-08-25 | 2015-11-11 | 辽宁中医药大学 | Integrated micro-fluidic chip for screening drug compatibility, as well as preparation method and application of integrated micro-fluidic chip |
| CN108679448A (en) * | 2018-06-02 | 2018-10-19 | 哈尔滨工业大学 | Microfluid flow on-line control device and detection method |
Non-Patent Citations (1)
| Title |
|---|
| 王琪民: "《微型机械导论》", 31 March 2003, 中国科学技术大学出版社 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114512179A (en) * | 2022-02-02 | 2022-05-17 | 上海图灵智算量子科技有限公司 | Quantum mutual learning mechanism-based binding energy prediction method |
| CN114512179B (en) * | 2022-02-02 | 2024-09-24 | 上海图灵智算量子科技有限公司 | Combination energy prediction method based on quantum mutual learning mechanism |
| CN114471762A (en) * | 2022-02-22 | 2022-05-13 | 西北大学 | Design and implementation method of micro-fluidic chip capable of controlling ultralow flow velocity |
| AU2022279500B1 (en) * | 2022-12-01 | 2023-11-23 | Provectus IP Pty Ltd | Automated cell analyser and method |
| WO2024112995A1 (en) * | 2022-12-01 | 2024-06-06 | Provectus IP Pty Ltd | Automated cell analyser and method |
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